The enormous size of microbial populations has proved to be a great asset in a variety of studies, but only because it is possible to effectively select certain kinds of rare gene type or mutant microorganisms. Mutant varieties of a single strain of microorganism (procaryotic, eucaryotic or viral) have classically been isolated by a variety of methods including positive cell "selection" and differential "screening".
Selection is used to isolate mutant varieties of microorganisms when a genetic alteration provides the microorganism with a positive growth advantage over its parental population. For example, acquisition of antibiotic resistance can be used to select such mutants on a nutrient agar surface containing the antibiotic. Another example is the acquisition of a biosynthetic gene enabling the organism to grow in a culture medium that would not otherwise support growth. There are, however, other genetic alterations such as additions, substitutions or deletions of the microorganism's genome which affect the primary or secondary metabolism of the microorganism, causing a small change or negative change (decrease) in the rate of growth. Such alterations may result in a beneficial increase or decrease in the synthesis or the breakdown of chosen biochemicals. Under such circumstances, screening must generally be utilized to isolate the mutant colony. Screening may involve examination of tens of thousands of individual colonies to determine the presence of mutants. Replica plating is one such screening technique. In general, it can be said that screening techniques, although highly effective in achieving the desired result, are labor and material-intensive requiring examination of many individual colonies usually in petri dishes; replica plating and tedious visual comparison of petri dish pairs are required as well as relatively large amounts of selective and/or restrictive materials which serve to differentiate the mutant from its parent.
Recently, a technology has emerged which provides methods of encapsulating biological material such as living tissue, individual cells, viruses, and biological macromolecules within a semipermeable membrane. The basic approach in this technique involves suspending the biological material to be encapsulated in a physiologically compatible medium containing a water soluble substance that can be made insoluble in water, that is, a gel, to provide a temporary environment for the biological material. The medium is formed into droplets containing the tissue and gelled by changing any one of the variety of ambient conditions. These temporary capsules are then subjected to a treatment which results in the production of membranes with a desired permeability (including impermeable membranes). One such technique, is exemplified in U.S. Pat. No. 4,352,883 entitled "Encapsulation of Biological Material", the disclosure of which is incorporated herein by reference.
A description of a technique for separating cells having desired properties from a larger population is found in U.S. Pat. No. 4,401,755 entitled "Process for Measuring Microbiologically Active Material" which discloses a method for measuring an unknown quantity of microbiologically active material utilizing a microencapsulation techniques similar to U.S. Pat. No. 4,352,883. The disclosure of U.S. Pat. No. 4,401,755 is also incorporated herein by reference. After preparing a suspension of gel microdroplets, the suspension is processed in an apparatus having the capability of sensing a physical characteristic of individual gel microdroplets to determine the presence or absence of a desired physical characteristic of the biological material in such a droplet.
Microencapsulation technology, as described in the above referenced patents, provides the potential for solving a variety of problems including the labor and cost excesses of prior art mutant microorganism isolation techniques. It is apparent that a need to develop new isolation techniques exists which will reduce the costs and time spent in selection and screening processes used to isolate mutants from their respective parent populations.